Barley possesses the highest level of salt tolerance among cereals, thus it is used as an important species to investigate mechanisms involved in salt tolerance. The present investigation was conducted to study the effect of salt stress by different NaCl concentrations (0, 9000, 12000, and 15000 ppm) on proline production and it was found that there is a positive correlation between proline production and salinity. Also study the effect of salinity on BADH-1 gene expression by Real-Time PCR,where this gene was performed and it was aligned in GenBank. The sequencing was translated into amino acid sequences and these amino acid sequences were aligned in GenBank. Finally, the secondary structure of BADH-1 enzyme was determined from the most tolerant barley (Giza 2000 and Wadi Sedr) and sensitive barley Giza 129 cultivars, then sequencing translated into amino acid sequences and these sequences were aligned in GenBank. Finally, the secondary structure of BADH-1 enzyme was determined. These isolated genes were submitted in GenBank under accession numbers KX433169, KX342849, KX342850, KX342851, KX433170, and KX433171.